The development and commercialization of many processes in the fields of medicine, chemistry, and agriculture require the use of fermentation devices or “bioreactors.” As used herein, the term “fermentation” means a process for the production of a product by culturing cells or microorganisms, the process of culturing cells or microorganisms, or a process for the bioconversion of one material to another. In some bioprocesses, the organisms themselves are the desired product.
Cell culturing, for example, is often carried out in vessels that permit the contacting of cells with nutritive media and oxygen. In industrial applications, such processes are often carried out in very large vessels, often far greater than 50 liters in capacity. During research and development, however, it is generally desirable to test such processes on a much smaller scale. Historically, fermentation devices and bioreactors with volumes of 50 liters or less have suffered from various deficiencies.
Many problems with existing devices lie with the mixing mechanisms employed. Some processes use enzymes immobilized on the surfaces of particles within a liquid medium. As a result, most of the enzymatic activity is limited to the surfaces of the particles. Any method of mixing the liquid medium that causes abrasion of the particles will necessarily reduce enzymatic activity. Similar damage can be caused to cells or microorganisms within a liquid medium.
Magnetic stirrers, for example, are inapplicable to some processes, including the culturing of cells or microorganisms, because of the tendency of the magnetic stirrer, that necessarily contacts an interior surface of the vessel, to damage delicate components, such as living cells and microorganisms, that become trapped between the magnetic impeller and the vessel surface. Attempts have been made to alleviate this disadvantage through the use of super conductive materials. TC Tech Corp. (www.tc-tech.com), for example, markets a mixing device wherein a disposable impeller is levitated above the vessel's bottom, thereby eliminating the potential for entrapment of cells or microorganisms between the impeller and the vessel wall. Such devices are, however, expensive to use, due to the desirability of operating the magnets at super conducting temperatures.
U.S. Patent Application Publication No. 2003/0008389 to Carll describes a disposable cell culture vessel with a hollow sleeve in its interior, into which is placed a magnetic stirrer. In some embodiments, the sleeve is fitted with a flexible blade. Such a device also reduces or eliminates the tendency of magnetic mixers to damage delicate components. However, due in part to the fact that the mixing action of the device is provided by the simple rotation of a magnetic bar, the device is incapable of providing greater agitation or aeration of the liquid medium. Rather, the placement of a magnet within the hollow sleeve allows the gentle rotation of the impeller and the subsequent undulation of the flexible blades when an adjustable magnetic force, such as a magnetic stir plate, is applied to the vessel. This creates a gentle stirring of the cells, which keeps the cells in suspension and prevents the cells from shearing. Where more vigorous agitation or greater aeration of the liquid medium is needed, such a device is inadequate.
Other devices use blades or similar mechanisms to mix their liquid contents. U.S. Pat. No. 3,468,520 to Duryea et al., for example, describes a paddle-like mechanism residing within a bottle, which is designed to agitate a suspension of cells. Such devices, however, require the introduction of a foreign object, in the form of the mixing mechanism, into the liquid medium. This greatly increases the possibility of contamination of the medium by substances or organisms residing on the mixing mechanism. Avoidance of such contamination requires thorough cleaning and sterilization of the mixing mechanism before each use, which can greatly increase not only the burden and expense of using such devices, but also the level of technical experience required by its users.
Others have attempted to integrate the mixing mechanism into the vessel itself. U.S. Pat. No. 3,432,149 to Stalberg et al., for example, describes an apparatus for stirring a liquid having internal wings, wherein rotation of the device along its longitudinal axis exerts a dragging action on the liquid. However, such a device is capable of exerting a dragging action on only a small portion of the liquid. “The height of the liquid-dragging part of the vessel is at the most half of the intended liquid level, suitably no more than one-third thereof and preferably about one-fourth thereof.” Col. 2, lines 43-46. In addition, such a device is incapable of aerating the liquid by, for example, projecting a portion of the liquid above the level of the standing liquid, thereby creating turbulence between the surface of the liquid and a gaseous layer above it.
Attempts have been made to eliminate the need for internal mixing mechanisms altogether. U.S. Pat. No. 4,373,029 to Nees, and U.S. Pat. No. 3,540,700 to Freedman et al., for example, describe devices for pivotally rotating vessels containing cells and a nutrient medium. There is a limit, however, to the degree of mixing attainable with such devices. For example, Nees notes that “acceleration magnitudes are essentially determined only by the gravity of the micro carrier in the earth's gravitational field, reduced by the viscosity of the nutrient solution.” Col. 2, lines 11-14. Thus, for processes requiring a greater degree of mixing or agitation, including, for example, processes requiring greater aeration of the liquid medium, such devices are not useful.
Another method by Wave Biotech has no internal mixers and uses a flexible vessel that is pleated. The action resembles that of the bellows of an accordion. A portion of the vessel is compressed and released such that fluid in the pleats must squirt into the main bulk of the fluid to cause agitation and mixing. The pleated vessel is quite expensive to construct and cannot be scaled up to large sizes. Furthermore, the mixing is poorly related to that in commercial fermenters thus making this agitation method of questionable use when generating data for scale up.
A need exists, therefore, for a device that avoids the above described limitations. Specifically, there is a need for a fermentation chamber and mixing apparatus that (1) will not damage delicate components of the liquid medium, such as living cells and microorganisms, (2) can provide sufficient agitation of the liquid medium to ensure proper mixing and/or aeration, (3) is inexpensive to produce and use.